Methods for producing cultured pearls in conch and other gastropods

ABSTRACT

The subject invention provides methods for producing cultured pearls in gastropods, such as the queen conch,  Strombus gigas , and other gastropod species (e.g., families Strombidae, Haliotididae and Volutidae), with high success rates. The subject invention also provides cultured pearls produced by the methods as described herein.

CROSS-REFERENCE TO A RELATED APPLICATION

This application is a continuation of co-pending application Ser. No.12/751,316, filed Mar. 31, 2010, which claims the benefit of U.S.Provisional Application Ser. No. 61/165,157, filed Mar. 31, 2009, whichare incorporated herein by reference in their entirety.

BACKGROUND OF INVENTION

A pearl is formed as a result of a defense mechanism against foreignirritants by mollusks. For many species, following the intrusion of theforeign irritant, the mantle tissue of the mollusk secretes aragonite ora mixture of aragonite and calcite (both crystalline forms of calciumcarbonate) and conchiolin (a horn-like protein) to coat the irritant.The combination of aragonite and conchiolin is called nacre, ormother-of-pearl. The secretion is repeated many times and the irritantis then covered by several layers of nacre, forming a pearl.Accordingly, pearls produced by the nacre secretion process are callednacreous pearls.

In addition, there are also non-nacreous pearls (sometimes referred toas “calcareous concretions”), that are porcellaneous (i.e., have alow-luster, ceramic-like surface). Many pearls of this type arenon-attractive and valueless but among them, pearls in species such asconch (Strombus gigas), and various scallop species are still ofinterest.

A conch pearl is produced by the conch family Strombidae. Conch pearlsdisplay various colors that range from white to brown with a widevariation in yellow, pink and orange hues (Fritsh and Misiorowski, 1987;Federman and Bari, 2007). For example, the queen conch, Strombus gigas,naturally produces valuable pearls that can exhibit an attractive pinkcolor (Acosta-Salmón and Davis, 2007). The conch pearl frequentlyexhibits a characteristic “flame” structure or pattern on its surface.This flame effect, also called “chatoyancy,” is caused by fibrousprismatic crystals perpendicularly aligned to the surface of the pearl(Federman and Bari, 2007). Unfortunately, natural conch pearl productsare lacking around the world because of their rarity. Their desirabilitywarrants the need for cultured conch pearls.

There are two main techniques for the production of cultured pearls. Oneis the technique used in marine pearl oysters, developed and refined bythe Japanese in the early 1900s, in which a nucleus (or bead) and apiece of mantle tissue from a donor pearl oyster are implanted into thegonad of a recipient pearl oyster, a process known as “grafting,”“beading” or “seeding” (Acosta-Salmón et al., 2004; CIBJO, 2006).Generally only one or two nuclei can be implanted at one time, dependingon the pearl oyster species and the grafting technician.

The second technique is used in freshwater mussels. This technique hasmany variations and has been continuously developing (Fiske andShepherd, 2007). Briefly, it involves grafting pieces of mantle tissuefrom a donor mussel into the mantle of a recipient mussel (Dan andRuobo, 2003; Fiske and Shepherd, 2007); this process is frequentlycalled “tissue nucleation,” although this term is incorrect because themantle tissue is never the nucleus in a non-beaded cultured pearl.Grafting success rates for cultured pearl production vary widely. Forexample, mortality of freshwater mussels at the first pearl harvest isaround 90% (Fiske and Shepherd, 2007). On the other hand, after graftingthe blacklip pearl oyster, Pinctada margaritifera, 10% of the oystersdied and a further 20% rejected the nucleus (Ellis and Haws, 1999).Mortalities between 2 and 24% and bead rejections between 9 and 16% wereobserved in P. margaritifera subjected to different post graftingtreatments (Norton et al., 2000).

U.S. Pat. No. 3,871,333 discloses a method of producing cultured pearlsin abalones by perforating a hole through the shell of the abalone,depositing a nucleus bead on the reproductive organ of the abalonethrough the hole, covering the hole, and raising the treated abaloneuntil pearls are formed.

EP Patent Application No. 1,084,615 discloses a nucleus for producingblister or mabé pearls in mollusks. The nucleus is generally bell-shapedwith a dome and a peripheral edge portion. The surface of the peripheraledge portion and some or all of the dome includes one or more stepswhich start at, or adjacent to, the outer edge. The steps assist byproviding a key for the deposition of nacre. The nucleus may be securedto the shell of the mollusk in a variety of ways.

U.S. Pat. No. 5,347,951 discloses a process for nucleating pearls inshell-bearing mollusks by forming an opening in the shell of a hostmollusk in a region covering a soft tissue, providing a pearl nucleushaving a first portion around which nacre forms and a second portionsecurely connected to the first portion and having a region larger thanthe size of the opening, and manually inserting the nucleus through theopening to an implanted position.

PCT Application No. PCT/NZ98/00167 discloses a nucleus for theproduction of half or mabe pearls in mollusks. The nucleus comprises afirst portion having an external surface adapted to define the shape ofthe half or mabe pearl to be produced and a second portion that definesa bridge between the first portion and the shell of the host mollusk.The half or mabe pearls so produced have a substantially even crosssection of nacre formed over the nucleus, and a generally reduced timeis required for the formation of the pearl.

Published U.S. Patent Application No. 2004/0112086 discloses a method ofproducing pearls including inserting a nucleus into a mollusk able toproduce a pearl, incubating the nucleus within the mollusk, and removinga portion of the nacre coating, thereby exposing a portion of thenucleus.

The techniques described in the above disclosures all have certaindrawbacks, such as the necessity for perforating a hole in the shell ata location covering a soft tissue, which is often difficult and causeshigh mortality. Thus, although queen conch aquaculture methods are wellestablished (Davis and Shawl, 2005), attempts to develop techniques toproduce cultured pearls in this species have been unsuccessful to date(Federman and Bari, 2007).

There is a considerable need for a method of commercially culturingconch (family Strombidae) pearls and pearls from other gastropods (e.g.,families Haliotididae and Volutidae). The subject invention providesmethods for culturing non-beaded and beaded conch pearls with successrates (retention and survival) comparable with those achieved incommercial oyster pearl operations.

BRIEF SUMMARY

The subject invention provides methods for the production of culturedpearls in the queen conch, Strombus gigas, and other gastropod species(e.g., families Strombidae, Haliotididae and Volutidae).

In a preferred embodiment, the method of the subject invention comprisespretreating a recipient conch with a relaxant, cutting an incision intothe recipient conch; cutting a subepithelial canal through the incision,inserting a mantle tissue piece in the subepithelial canal, optionallyinserting a nucleus in the subepithelial canal, culturing the recipientconch to obtain a cultured pearl, and harvesting the cultured pearl.

The incision may be performed on the skin of any part of the body insidethe shell of a recipient gastropod. Preferably the incision is made onthe skin of the foot and the subepithelial canal is cut into the softtissue of the conch.

In one embodiment, beaded pearls are formed by inserting both the mantletissue piece and the nucleus in the subepithelial canal.

In another embodiment, non-beaded pearls can be cultured by insertingonly the mantle tissue in the canal.

The subject invention also provides cultured pearls produced by themethod as described herein. In one embodiment, the subject inventionprovides nucleated cultured pearls, wherein the pearl comprises an innernucleus and outer non-nacreous, calcareous layers. In anotherembodiment, the subject invention provides non-nucleated culturedpearls, wherein the pearl comprises an inner cavity and outernon-nacreous, calcareous layers. In a preferred embodiment, the pearl iscultured in Strombus gigas.

DETAILED DISCLOSURE

The subject invention provides methods for the production of culturedpearls in the queen conch, Strombus gigas, and other gastropod species(e.g., families Strombidae, Haliotididae and Volutidae). The subjectinvention also provides nucleated and non-nucleated pearls cultured inthe queen conch, Strombus gigas, and other gastropod species.

In a preferred embodiment, the method of the subject invention comprisespretreating a recipient conch with a relaxant, cutting an incision intothe recipient conch; cutting a subepithelial canal through the incision,inserting a mantle tissue piece in the subepithelial canal, optionallywidening the incision and inserting a nucleus in the subepithelialcanal, culturing the recipient conch to obtain a cultured pearl, andharvesting the cultured pearl. In a preferred embodiment, the shell ofthe conch is not perforated.

Preferably the incision is made on the skin of the foot and thesubepithelial canal is cut into the soft tissue of the conch.

In one embodiment of the subject invention, beaded pearls are formed byinserting both the mantle tissue piece and the nucleus in thesubepithelial canal.

In another embodiment, non-beaded pearls can be cultured by insertingonly the mantle tissue in the canal.

The subject invention also provides cultured pearls produced by themethod as described herein. In one embodiment, the subject inventionprovides nucleated cultured pearls, wherein the pearl comprises an innernucleus and outer non-nacreous, calcareous layers. In anotherembodiment, the subject invention provides non-nucleated culturedpearls, wherein the pearl comprises an inner cavity and outernon-nacreous, calcareous layers. In a preferred embodiment, the pearl iscultured in Strombus gigas.

In one embodiment, the cultured pearls comprise a taggant that makes itpossible to identify pearls produced according to the subject method.The taggant can be any detectable molecule and/or source of radiation.The taggant may be associated with a nucleus used in the method of thesubject invention. In preferred embodiments, the nucleus is one that isnot found in pearls produced naturally in conch.

According to one aspect of the subject invention there is provided amethod of growing pearls, comprising the steps of:

pretreating a recipient gastropod by a relaxant;

cutting an incision into the recipient gastropod in the skin of thefoot;

cutting a subepithelial canal through the incision;

inserting a mantle tissue piece and/or a nucleus from a donor in thesubepithelial canal;

culturing the recipient gastropod to obtain a cultured pearl; and

harvesting the cultured pearl.

Preferably the recipient is a queen conch, Strombus gigas. However it isenvisaged that any conch (e.g., family Strombidae) or similar gastropodspecies (e.g., Melo melo and Haliotis sp. and other species in thefamilies Haliotididae and Volutidae) can be used as the recipientaccording to the subject invention.

The mantle donor is preferably a mollusk. Preferably a healthy conch isused as the donor conch. Preferred donor species include any conch(e.g., family Strombidae) or similar gastropod species (e.g., Melo meloand Haliotis sp. and other species in the families Haliotididae andVolutidae). However it is envisaged that any mollusk species, such asfor example, gastropods, saltwater mollusks, freshwater mussels,abalones, can be used as the mantle donor according to the subjectinvention. The mantle donor conch is selected following similar criteriato that used for pearl oyster mantle donors. For example, a healthyconch has a healthy shell and well defined (e.g., pink) coloration inthe interior of the shell.

In a preferred embodiment of the subject invention, the cutout mantletissue piece has a generally rectangular shape and is, for example,about 2 mm to about 5 mm in length and/or width. Alternatively, thecutout mantle tissue piece has a size of about 2.5 mm to about 4.5 mm,or about 3.5 mm to about 4 mm. For a queen conch, the preferred size ofthe piece is between about 2-4 mm.

In a preferred embodiment of the method of the subject invention arelaxant is used for relaxation of the recipient conch. Examples of thechemical relaxant include, but are not limited to, chloral hydrate,2-phenoxyethanol, benzocaine, magnesium chloride, menthol and anychemical combination thereof. It is preferred that the relaxants anddose thereof have rapid response, are non-toxic, and allow for completerecovery of the conch. For example, the recipient queen conch can berelaxed by about 30 g/L of magnesium chloride for at least about 20minutes.

The incision may be performed on the skin of any part of the body insidethe shell of a recipient gastropod. Preferably the incision is made onthe skin of the foot and the subepithelial canal is cut into the softtissue of the conch.

In general, up to about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 incisions maybe performed on each recipient conch. In certain embodiments, up toabout 15, 20, 25, 30, 35, or 40 incisions may be performed on eachrecipient gastropod. The optimal number of incisions per recipient mayvary, depending on the size of the recipient, the size and shape ofnucleus, the size and shape of mantle tissue, and/or the positioning ofthe mantle tissue piece and the nucleus in the subepithelial canal, allof which can be readily determined by those skilled in the art havingthe benefit of the teachings of the present invention.

The subepithelial canal, at the end of which the nucleus and/or themantle tissue piece is inserted, is cut through the incision. In oneembodiment, a cut is made between the muscle and skin.

The size of the incision and/or the subepithelial canal may be of anywidth or depth suitable for inserting the mantle tissue piece and/or thenucleus into the recipient gastropod species. For the production ofnon-beaded pearls, the depth and/or width of the incision and/or thesubepithelial canal is typically up to about 3 mm, about 4 mm, about 5mm, about 6 mm, or about 7 mm. For production of beaded conch pearls,the depth and/or width of the subepithelial canal is typically up toabout 6 mm, about 7 mm, about 8 mm, about 9 mm, about 10 mm, about 13mm, about 15 mm, or about 18 mm.

In a preferred embodiment, the mantle tissue piece is inserted withshell-producing cells facing the incision.

In a preferred embodiment, the nucleus should be commercial grade, suchas for example, commercial grade nucleus made from freshwater mussels ofthe Mississippi River, similar to those used in the pearl oysterindustry.

In preferred embodiments, the nucleus is not sand, dirt, shell from therecipient, or other materials that act as the nucleus when pearls areproduced naturally in conch. In one embodiment, the nucleus is free ofharmful bacteria, or is even sterile.

Typically, the nucleus is spherical in shape, and has a diameter ofabout 5 mm to about 10 mm, although a nucleus of less than 5 mm orgreater than 10 mm may also be used. The size of the nucleus shouldallow insertion into the canal. Preferably the nucleus is gently pushedto maintain the normal state of the conch. Too much pressure may causethe nucleus to pierce the skin or body.

According to one aspect of the subject invention, there is provided amethod of growing beaded pearls, wherein both a nucleus and a mantletissue piece are inserted.

In a preferred embodiment, the subject invention uses a recipient conchthat is sufficiently large to receive the nucleus. The larger the conch,the larger the nucleus it can receive.

According to another aspect of the subject invention, there is alsoprovided a method of growing non-beaded pearls, wherein only a mantletissue piece per incision is inserted.

The subject invention also provides a pearl that has been cultured in aconch or other gastropod using a method of the subject invention. Thus,in one embodiment the subject invention provides pearls produced byinserting into a recipient conch a nucleus and/or a mantle tissue piecethrough an incision into a subepithelial canal.

The nucleus used according to the subject invention may be specificallydesigned so that the pearl produced by the method of the subjectinvention can be identified. Thus, the nucleus may have, for example, anidentifiable taggant. The taggant may be identifiable through physicalmeans, including shape, density, size, and the like; chemical means;and/or radiological means.

The cultured pearls may have a substantially spherical (beaded) shape; asubstantially symmetrical shape such as pear, oval, tear drop, or heart;or a baroque shape. The shape may be influenced by the physicalcharacteristics of the nucleus and the mantle tissue piece and theculture conditions of the cultured conch. The cultured pearls maydisplay various colors that range from white to brown with a widevariation in yellow, pink and orange hues.

The size and weight of the cultured pearl may vary, depending on thesize of the recipient, the size and shape of nucleus, the size and shapeof mantle tissue, and/or the culture time and condition. In certainembodiments, the cultured pearls of the subject invention have a size(length and/or diameter) of about 2 mm to about 15 mm, about 3 mm toabout 12 mm, about 4 mm to about 10 mm, or about 5 mm to about 7 mm indiameter. However, cultured pearls with a size of less than 2 mm orgreater than 15 mm may also be formed. In a specific embodiment, thecultured conch pearls have a size of about 2 mm to about 8 mm, or about3 mm to about 5 mm in diameter.

In certain embodiments, the cultured pearls of the subject inventionhave a weight of about 0.5 to about 7 carats, about 1 to about 5 carats,about 1.5 to about 3 carats, or about 2 to about 3 carats. However,cultured pearls with a weight of less than 0.5 carat or greater than 5carats may also be formed. In a specific embodiment, the cultured conchpearls have a weight of about 0.5 to about 5 carats, about 1 to about 4carats, about 1.5 to about 3 carats, or about 2 to about 3 carats.

The culture time may vary according to the conch and the culturecondition. Typically, it takes over one year of culture to produce queenconch non-beaded cultured pearls with a mean weight of 1-2 ct (1 ct=200mg) and about one year to produce beaded cultured pearls larger than 7mm in diameter (3-5 et) in the queen conch if beaded with 5.1 mm nuclei(1.7 bu).

The cultured pearls may comprise molecules associated with the recipientgastropod. These molecules may be, for example, conch (or othergastropod) protein and/or DNA. These molecules can be detected utilizingassays that are well known in the art. Advantageously, the nucleus ofthe cultured pearl is, preferably, different from the nucleus that wouldbe found in natural pearls.

Advantageously, pearls can be produced in large numbers according to thesubject invention. This unique process makes it possible, for the firsttime, to transport conch (or other gastropod) pearls in largequantities. Thus, in one embodiment, the subject invention provides acontainer having enclosed therein multiple conch (or other gastropod)pearls. Preferably the container has more than 5, 10, 15, 20, 25, 50,100, or even 500 pearls. Examples of containers include boxes, bags,sacs and the like. Preferably the container is man-made. The containerpreferably encloses a volume of less than 10, 8, 5, 3, 2, 1, 0.5, 0.2,0.1 or even 0.05 liters.

The cultured pearls of the present invention can be identified usingstandard identification techniques well known in the jewelry industry.For instance, conch pearls can be identified based on theirnon-nacreous, calcareous compositions and the characteristic “flamestructure” that is reminiscent of a fire burning on the surface. Inaddition, cultured pearls can be distinguished from natural pearls usingX-ray examination to unveil the internal growth structure. For instance,bead-nucleated cultured pearls typically have a solid nucleus in thecenter without any concentric growth ring, whereas natural pearls haveconcentric growth rings. Also, beadless, non-nucleated cultured pearlsmay have growth rings, but they also have a void, inner cavity. Thus,the type, quality, shape, size, and color of the cultured pearls can bereadily determined by those skilled in the jewelry industry.

Following are examples that illustrate procedures for practicing theinvention. These examples should not be construed as limiting.

EXAMPLE 1 Culturing Queen Conch, Strombus Gigas, Pearls

A medium sized queen conch was relaxed with 30 g of magnesium chloridefor at least 20 minutes. A large pearl oyster shell holder was used tohold the conch and a heavy vice-grip was used to clamp on the conch'soperculum and to gently pull the conch out of the shell. To obtainmantle tissue a pair of tissue forceps and a scalpel were used. Thecutout piece of tissue was then laid on a plastic cutting board and blotdried using a sponge or paper towel. The same scalpel was used to cutthe mantle tissue into small squares.

For the production of non-beaded cultured pearls, the scalpel was usedto cut a small incision into the tough skin on the conch foot. Once thisincision was made, the mantle or tissue inserter was used to cut asubepithelial canal into the conch soft tissues. After that, a piece ofmantle tissue was inserted using a tissue inserter. Due care to placethe tissue with the shell producing cells facing the incision, should betaken.

For the production of beaded cultured pearls, in addition to theprevious steps, a hook and a nucleus inserter can be used. After thesmall incision is made with the scalpel knife, a side knife can be usedto widen the incision and the canal. Using the guided scalpel, anincision is made into the conch soft tissues, carefully guiding thescalpel between the foot muscle and the foot skin. After the incision, apiece of mantle tissue can be inserted using a tissue inserter. Anucleus can then be inserted through the incision using a nucleusinserter. It is difficult to observe the piece of mantle tissue makingcontact with the nucleus. However, this technique has proven to be veryeffective. If applied correctly, this technique will provide almost 100%nucleus retention and 0% mortality 6 weeks after implant.

It should be understood that the examples and embodiments describedherein are for illustrative purposes only and that various modificationsor changes in light thereof will be suggested to persons skilled in theart and are to be included within the spirit and purview of thisapplication.

All patents, patent applications, provisional applications, andpublications referred to or cited herein are incorporated by referencein their entirety, including all figures and tables, to the extent theyare not inconsistent with the explicit teachings of this specification.

REFERENCES

-   Acosta-Salmón, H., Davis, M., 2007. Inducing relaxation in the queen    conch Strombus gigas (L.) for cultured pearl production. Aquaculture    262, 73-77.-   Acosta-Salmón, H., Martinez-Fernandez, E., Southgate, P. C., 2004. A    new approach to pearl oyster broodstock selection: can saibo donors    be used as future broodstock? Aquaculture 231, 205-214.-   CIBJO, 2006. The Pearl Book. Natural, Cultured and Imitation    Pearls—Terminology and Classification. CIBJO/Gem Materials. CIBJO 60    pp. http://www.gema.    com/information/CIBJO/CIBJO%20Pearl%20Book%20-%202006-1.pdf    (accessed on Feb. 15, 2008)-   Dan, H., Ruobo, G., 2003. Freshwater pearl culture in China. Global    Aquaculture Advocate 6(3), 50-51.-   Davis, M., Shawl, A. L., 2005. A guide for culturing queen conch.    Am. Fish. Soc. Symp. 46, 125-142.-   Ellis, S., Haws, M., 1999. Producing pearls using the black-lip    pearl oyster (Pinctada margaritifera). Aquafarmer Information Sheet.    Center for Tropical and Subtropical Aquaculture Special Publication    No 141. December 1999.-   Federman, D., Bari, H., 2007. The Pink Pearl. A Natural Treasure of    the Caribbean. Skira, Milan, Italy. 176 pp.-   Fiske, D., Shepherd, J., 2007. Continuity and change in Chinese    freswater pearl culture. Gems Gemmol. 43, 138-145.-   Fritsch, E., Misiorowski, E. B., 1987. The history and gemology of    queen conch ‘pearls’ Gems Gemmol. 23(3), 208-221.-   Norton, J. H., Lucas, J. S., Turner, I., Mayer, R. J., Newnham,    R., 2000. Approaches to improve cultured pearl formation in Pinctada    margaritifera through use of relaxation, antiseptic application and    incision closure during bead insertion.

We claim:
 1. A method of growing a pearl in a gastropod comprising thesteps of: a) pretreating a recipient gastropod with a relaxant; b)cutting an incision into the tissue of the recipient; c) cutting asubepithelial canal through the incision; d) inserting a mantle tissuepiece and/or a nucleus from a donor in the subepithelial canal; e)culturing the recipient gastropod to obtain a cultured pearl, and f)harvesting the cultured pearl.
 2. A cultured pearl, produced by a methodaccording to claim
 1. 3. The cultured pearl, according to claim 2,wherein the cultured pearl is nucleated.
 4. The cultured pearl,according to claim 2, wherein the cultured pearl is non-nucleated.
 5. Acultured pearl that comprises one or more molecules of gastropod origin,or which has been produced in a gastropod and has a nucleus that is notfound in natural gastropod pearls.
 6. The pearl, according to claim 5,which comprises DNA and/or protein of conch origin.
 7. The pearl,according to claim 5, comprising a nucleus comprising material from aMississippi River mollusk.
 8. The pearl, according to claim 5, whereinthe nucleus comprises a physical, chemical, or radiological taggant. 9.A container holding at least 5 pearls according to claim 5.